Sequence of the novel joints present in the amplified DNA of N-phosphonacetyl-L-aspartate resistant Drosophila cells: Implication on the mechanisms of amplification in these cells

Summary— We have previously shown the presence, in the amplified DNA of a Drosophila cell line resistant to N-phosphonacetyl-L-aspartate (PALA), of two units of 150 kb and 120 kb respectively duplicated and amplified. The two joints (J1 and J2) linking these units as well as their respective wild-type counterparts have been sequenced. Sequence analysis indicates that a region of the Drosophila genome which corresponds to the proximal boundary of the 150 kb unit is common to both joints. In addition to this common region, the J1 junction possesses a 26-nucleotide sequence belonging to the J2 junction. This indicates that the J2 junction was the first formed, and that J1, therefore, results from recombination between J2 and a region of the wild-type genome 120 kb distal to J2. Sequence analysis also reveals that the joints result from illegitimate recombination between unrelated regions. AT-rich sequences, strand bias composition and putative topoisomerase I and II sites were found in at least one of the two parental sequences involved in the formation of the joints. On the basis of these results we can hypothesize that after two illegitimate recombinations between sister chromatids, leading first to J2 and then to J1, the amplification may have arisen by a series of homologous (unequal crossing-over) or illegitimate recombinations, or by an intrachromosomal rolling circle.     

Effect of lead on the reaction of O-demethylase in p-nitro anisole in the cladoceran Moina macrocopa

The presence of the Mixed-Function Oxydase (MFO) system, in Moina macrocopa was detected through the transformation of p-nitroanisole to para-nitrophenol. The presence of this enzymatic system suggested that this cladoceran participates in the biotransformation of xenobiotics in aquatic ecosystems. This capacity, in conjunction with the aquatic flea's high tolerance to environmental stress, suggested that M. macrocopa could be used in bioremediation efforts to increase ecosystem health. The effects of lead on the MFO system in M. macrocopa, were also studied. Lead acted as an inhibitor of the enzymatic complex. Therefore, induction of MFO-activity as an early warning may not work in waterbodies affected by both inducers of MFO and inhibitors like lead.     

Characterization of Xanthomonas campestris Pathovars by rRNA Gene Restriction Patterns

Genomic DNA of 191 strains of the family Pseudomonadaceae, including 187 strains of the genus Xanthomonas, was cleaved by EcoRI endonuclease. After hybridization of Southern transfer blots with 2-acetylamino-fluorene-labelled Escherichia coli 16+23S rRNA probe, 27 different patterns were obtained. The strains are clearly distinguishable at the genus, species, and pathovar levels. The variability of the rRNA gene restriction patterns was determined for four pathovars of Xanthomonas campestris species. The 16 strains of X. campestris pv. begoniae analyzed gave only one pattern. The variability of rRNA gene restriction patterns of X. campestris pv. manihotis strains could be related to ecotypes. In contrast, the variability of patterns observed for X. campestris pv. malvacearum was not correlated with pathogenicity or with the geographical origins of the strains. The highest degree of variability of DNA fingerprints was observed within X. campestris pv. dieffenbachiae, which is pathogenic to several hosts of the Araceae family. In this case, variability was related to both host plant and pathogenicity.     

Localization of the lampbrush loop pair Nooses on the Y chromosome of Drosophila hydei by fluorescence in situ hybridization

We have used fluorescence in situ hybridization to map the positions of the different repetitive DNA sequences from the region forming the lampbrush loop pair Nooses on the Y chromosome of Drosophila hydei. This region harbours a megabase cluster of tandemly organized repeats of the Y-specific ay1 family and a megabase cluster of tandem repeats of the related Y-specific YsI family. In addition, ay1 repeats also occur in short blocks that are interspersed by other repetitive DNA sequences that we call Y-associated, since they have additional copies on other chromosomes. Using specific probes for ay1, YsI and Y-associated DNA sequences, we show that there is one large proximal cluster of YsI repeats and one, more distally located, large cluster of ay1 repeats. The Y-chromosomal copies of the Y-associated sequences are located in the most distal part of the ay1 cluster. This is consistent with the juxtaposition of ay1 and Y-associated sequences in more than 300 kb of cloned genomic DNA. Since both ay1 and Y-associated sequences have been shown to be transcribed in the Nooses, the lampbrush loop is formed in a distal region of the short arm of the Y chromosome, adjacent to the terminally located nucleolus organizer region. The clusters of homogeneous ay1 and YsI repeats are of no functional significance for the formation of the lampbrush loop.     

Iron content of sediment and phosphate adsorption properties

Phosphorus can occur in sediments in different forms and accordingly its availability varies. The distinction between the phosphorus fractions is made with two chemical extraction methods; an ammonium oxalate-oxalic acid extraction and an extraction according to Hieltjes & Lijklema (1980).The iron and aluminum liberated with the ammonium oxalate-oxalic acid extraction method is linearly correlated (r ² = 0.73) with the phosphorus liberated in the first two steps of the Hieltjes and Lijklema extraction by: P = 0.035 (Fe + Al) + 0.001 (P, Fe and Al in mmol g^–1).The iron and aluminum (hydr)oxides are very important fractions in the sediment adsorption capacity for phosphorus. The phosphorus sorption capacity (PSC) is 0.080 mol P (mol (Fe + Al))^–1 and the adsorption constant (k) is 11.9 µmol P l^–1. Here it is assumed that iron and aluminum (hydr)oxide have the same affinity for phosphorus.     

The genome of the non-cultured,bacterial-like organism associated with citrus greening disease contains thenusG-rplKAJL-rpoBC gene cluster and the gene for a bacteriophage type DNA polymerase

We have recently cloned three DNA fragments (In-2.6, In-1.0, and In-0.6) of the noncultured, bacterial-like organism (BLO) associated with citrus greening disease. Nucleotide sequence determination has shown that fragment In-2.6 is part of therplKAJL-rpoBC gene cluster, a well-known operon in eubacteria. The DNA fragment upstream of and partially overlapping with In-2.6 could be isolated and was shown to be thenusG gene. InEscherichia coli, nusG is also immediately upstream ofrplKAJL-rpoBC. Fragment In-1.0 carries the gene for a bacteriophage type DNA polymerase. Fragment In-0.6 could not be identified.When In-2.6 was used, at high stringency, as a probe to detect greening BLO strains in infected plants, hybridization was obtained with all Asian strains tested, but not with the African strain examined. At lower stringencies, In-2.6 was able to detect also the African strain. The implications of these reults in the taxonomical position of the greening BLO are discussed.     

Cyclitol metabolism is a central feature of Burkholderia leaf symbionts

The symbioses between plants of the Rubiaceae and Primulaceae families with Burkholderia bacteria represent unique and intimate plant–bacterial relationships. Many of these interactions have been identified through PCR-dependent typing methods, but there is little information available about their functional and ecological roles. We assembled 17 new endophyte genomes representing endophytes from 13 plant species, including those of two previously unknown associations. Genomes of leaf endophytes belonging to Burkholderia s.l. show extensive signs of genome reduction, albeit to varying degrees. Except for one endophyte, none of the bacterial symbionts could be isolated on standard microbiological media. Despite their taxonomic diversity, all endophyte genomes contained gene clusters linked to the production of specialized metabolites, including genes linked to cyclitol sugar analog metabolism and in one instance non-ribosomal peptide synthesis. These genes and gene clusters are unique within Burkholderia s.l. and are likely horizontally acquired. We propose that the acquisition of secondary metabolite gene clusters through horizontal gene transfer is a prerequisite for the evolution of a stable association between these endophytes and their hosts.     

The differential effect of N6-benzyl-adenine and N6-(Δ2-isopentenyl)-adenine on in vitro propagation of Paeonia suffruticosa Andr. is correlated with different hormone contents

Summary N⁶-benzyl-adenine (BA) enhanced phyllogenesis and axillary bud development of Paeonia suffruticosa during in vitro culture allowing good propagation while N⁶-(²isopentenyl)adenine (iP) did not. During the first five days of culture, the mitotic activity of BA-treated explants was higher than in the iP-treated ones. High BA levels were detected in the BA-treated explants, and this was correlated with the absence of or the low indole-3-acetic acid (IAA) content. The low iP levels measured in iP-treated explants were correlated with high endogenous IAA content; the new cytokinin / auxin ratio could explain the lack of axillary buds and the development of only one leaf. Abscisic acid (ABA) was detected neither in the controls nor in the cytokinin-treated explants during the first week. However, intensive restoration of ABA accumulation was observed in controls from the third week onwards. Both BA and iP-treated explants accumulated less ABA than the controls but this hormone appeared later in the BA-treated explants than in the iP-treated ones.Abbreviations ABA abscisic acid - BA N⁶-benzyl-adenine - BHT butyl-hydroxy-toluene - ELISA enzyme linked immunosorbent assay - FM fresh mass - HPLC high performance liquid chromatography - IAA indole-3-acetic acid - iP N⁶-(²-isopentenyl)adenine - MI mitotic index - 9RBA 9-ß-D-ribofuranosyl-BA - 9RiP 9-ß-Dribofuranosyl-iP - 9RZ 9-ß-D-ribofuranosyl-zeatin - Z zeatin     

A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex-determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.